Fig. 1
IKK2/NF-κB activation in oligodendrocytes induces an inflammatory phenotype in the CNS. a Immunoblot analysis of extracts from cortex and spinal cord tissue from control and IKK2-CAPLP−CreERT2 mice 3 weeks post induction (wpi). Strong expression of IKK2 and eGFP demonstrate tamoxifen-dependent transgene activation in IKK2-CAPLP−CreERT2 mice. High levels of phosphorylated p65 (p-p65) validate functional activation of NF-κB signaling upon transgene expression (n = 3). b Gene expression analysis of Ccl2 as an inflammatory marker and NF-κB target gene in tissue extracts from cortex, corpus callosum, cerebellum and spinal cord proves functionality of the transgenic system and depicts the inflammatory milieu within the CNS (n = 3–4). c and d Immunofluorescent staining and quantification of Iba1 reveals persistent microgliosis in the CC (3 wpi – Control (n = 4): 67 ± 6 cells/mm2, IKK2-CAPLP−CreERT2 (n = 4): 271 ± 64 cells/mm2, 20 wpi—Control (n = 6): 50 ± 7 cells/mm2, IKK2-CAPLP−CreERT2 (n = 5): 247 ± 31 cells/mm2). e and f Immunofluorescent staining and quantification of GFAP + astrocytes identifies an increased number of GFAP + astrocytes in the CC at 3 wpi that declines at 20 wpi (3 wpi – Control (n = 4): 3.50% ± 0.34 fluorescent area, IKK2-CAPLP−CreERT2 (n = 4): 19.62% ± 2.13 fluorescent area; 20wpi – Control (n = 6): 1.67% ± 0.53 fluorescent area, IKK2-CAPLP−CreERT2 (n = 5): 9.50% ± 2.53 fluorescent area). g and h Immunohistochemical analysis of oligodendrocytes (OLs) 3 and 20 wpi shows a significant increase in the overall CC1 + cell number in the corpus callosum (CC) of IKK2-CAPLP−CreERT2 animals at 20 wpi with a constant number of GFP + CC1 + cells (3 wpi—Control (n = 3): 5997 ± 398 cells/mm2, IKK2-CAPLP−CreERT2 (n = 3): 4979 ± 685 cells/mm2; 20 wpi – Control (n = 3): 7123 ± 147 cells/mm2, IKK2-CAPLP−CreERT2 (n = 4): 10,484 ± 1014 cells/mm2; GFP + cells- IKK2-CAPLP−CreERT2: 3 wpi: 3141 ± 575 cells/mm2, 20 wpi: 3883 ± 434 cells/mm2). i and j Immunohistochemical analysis of NG2 glia at 3 and 20 wpi reveals higher numbers of progenitor cells in IKK2-CAPLP−CreERT2 mice. White Arrows indicate NG2 glia. (3 wpi—Control (n = 3): 268 ± 17 cells/mm2, IKK2-CAPLP−CreERT2 (n = 3): 385 ± 20 cells/mm2; 20 wpi – Control (n = 3): 209 ± 8 cells/mm2, IKK2-CAPLP−CreERT2 (n = 4): 304 ± 15 cells/mm2). Data are shown as mean ± SEM. Grey bars, control; red bars, IKK2-CAPLP−CreERT2. Statistical analysis: One-way or Two-way ANOVA multiple comparison test followed by Bonferroni´s post hoc test (*p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 NS, non-significant (p > 0.05))